IMMOBILIZATION AND SOME BIOTECHNOLOGICAL APPLICATIONS OF CHITOSANASE FROM MONASCUS PURPUREUS

Polyamines induced chitosanase (Chsn, EC: 3.2.1.132) production by M. purpureus with increasing their concentrations. Polyamines could be arranged according to Chsn activity as agmatine < putrescine < spermine < spermidine < cadaverine. Chsn was purified from M. purpureus with specific activity of 35 units mg-1protein and 9.02-fold of purification. Chitosan 100 % of degree of acetylation (DA) as a substrate achieved maximum activity. The molecular weight of purified Chsn was 66 KDa. Km and Vmax for free Chsn were 1.4 mg/ml and 200 units mg-1 protein, respectively. However, Km and Vmax of immobilized Chsn on Ca-alginate were 2.5 mg/ml and 500 units mg-1 protein, respectively. The optimum pH values of free and immobilized enzymes on Ca-alginate were 6 and 7, respectively. The optimal temperatures were 40 °C and 50 °C for the two forms of the enzyme in the same order. Immobilized Chsn expressed appreciable heat stability at 60, 70 and 80°C. Kd values for immobilized Chsn were 10 x 10-2, 12.6 x 10-2 and 13.7 x 10-2. T0.5 values were 6.93, 5.50 and 5.01 min at 60 °C, 70 °C and 80 °C, respectively. It was observed that the inhibitory effect of chelating agents such as ethylenediaminetetraacetic acid (EDTA), o-phenanthroline, 8-Quinolinol, 8-Hydroxy-5-quinoline sulfonic acid (8-H5QS) and ?,?`- dipyridyl was less pronounced in the case of the immobilized enzyme and this give immobilized Chsn a great advantage. Chsn reaction product chitooligosacharides (CHOS) have antioxidant activity and can eliminate undesirable effects of reactive oxygen species in foods and scavenge some free radicals such as hydrogen peroxide (H2O2). Immobilized M. purpureus Chsn acts as strong inhibitor for chitinase and this plays an important role in malaria and asthma therapy.